Project Summary Metastasis, the dissemination of cells from the primary tumor, is the leading cause of death in cancer patients. For metastatic breast cancer, chemotherapy remains the standard of care. While the benefits of chemotherapy for the treatment of metastatic disease have been well documented, over 50% of triple-negative breast cancer patients become resistant to chemotherapy. Increasing our understanding of the mechanisms driving metastasis and drug resistance will help identify biomarkers that can be used to predict them, and characterize targetable signaling pathways to that can be used to prevent them. It is now well appreciated that the tumor microenvironment can contribute to tumor progression and metastasis. The extracellular matrix (ECM), forms a complex scaffold of proteins that provides both structure and signals to the tumor cells. However, its role in driving invasion and metastasis and how it could affect response to chemotherapy remains unexplored. The main goal of this proposal is to investigate the role of the ECM in metastasis and chemo-resistance, using an interdisciplinary approach that will combine cell biology, intravital imaging, systems biology and implantable devices. First, we have shown that gradients of fibronectin can promote directional motility of tumor cells, a process important for metastasis. When highly metastatic tumor cells are subject to gradients of EGF and fibronectin simultaneously, they will invade even more than with each cue alone. The goal of Aim 1 is to investigate the contribution of ECM versus growth factor cues during local invasion and metastatic colonization. Second, preliminary investigation into the effect of chemotherapeutic drugs on highly invasive tumor cells revealed that they can have differential effects on cell proliferation and migration, and can also regulate ECM sensitivity and organization. In Aim 2, I will identify ECM combinations that affect cell growth and migration in response to clinically relevant chemotherapy regimens and perform a CRISPR knockout screen of ECM-regulator proteins to identify novel genes regulating chemo-sensitivity. I also propose an extensive training program that will support my transition to independence. The research environment provided by the Koch Institute at MIT is outstanding, and offers unequaled opportunities for scientific discussion, collaboration between biologists and engineers, and career development. I have assembled an exceptional team to help me achieve my goals: Prof. Frank Gertler, expert in cell motility, and Prof. Michael Hemann, expert in studying mechanisms of drug resistance will be my mentors. Prof. Richard Hynes, a pioneer in ECM research, and Prof. Doug Lauffenburger, leader in cancer systems biology, will be collaborators and members of my mentoring committee. My training will also involve mentoring students, attending and presenting my work at meetings to become an active member of the ECM and cancer communities, and science outreach. Together, the proposed studies and career development training will ensure I achieve my goal of establishing a successful, independently-funded lab at a major university.